Peptide-containing α-ketoamide cysteine and serine protease inhibitors

ABSTRACT

This invention relates to peptide-containing α-ketoamide inhibitors of cysteine and serine proteases, methods for making these compounds, and methods for using the same.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a divisional of application Ser. No. 09/166,808,filed Oct. 6, 1998, the contents of which are incorporated herein in itsentirety which is a claims benefit of U.S. Provisional Application Ser.No. 60/061,309, filed Oct. 7, 1997, the disclosure of which is herebyincorporated herein by reference in its entirety.

FIELD OF THE INVENTION

This invention relates to peptide-containing α-ketoamide inhibitors ofcysteine and serine proteases, methods for making these compounds, andmethods for using the same.

BACKGROUND OF THE INVENTION

Numerous cysteine and serine proteases have been identified in humantissues. A “protease” is an enzyme which degrades proteins into smallercomponents (peptides). The terms “cysteine proteasell and “serineprotease” refer to proteases which are distinguished by the presencetherein of a cysteine or serine residue which plays a critical role inthe catalytic process. Mammalian systems, including humans, normallydegrade and process proteins via a variety of enzymes including cysteineand serine proteases. However, when present at elevated levels or whenabnormally activated, cysteine and serine proteases may be involved inpathophysiological processes.

For example, calcium-activated neutral proteases (“calpains”) comprise afamily of intracellular cysteine proteases which are ubiquitouslyexpressed in mammalian tissues. Two major calpains have been identified;calpain I and calpain II. While calpain II is the predominant form inmany tissues, calpain I is thought to be the predominant form inpathological conditions of nerve tissues. The calpain family of cysteineproteases has been implicated in many diseases and disorders, includingneurodegeneration, stroke, Alzheimer's, amyotrophy, motor neuron damage,acute central nervous system injury, muscular dystrophy, boneresorption, platelet aggregation, cataracts and inflammation. Calpain Ihas been implicated in excitatory amino-acid induced neurotoxicitydisorders including ischemia, hypoglycemia, Huntington's Disease, andepilepsy. The lysosomal cysteine protease cathepsin B has beenimplicated in the following disorders: arthritis, inflammation,myocardial infarction, tumor metastasis, and muscular dystrophy. Otherlysosomal cysteine proteases include cathepsins C, H, L and S.Interleukin-1β converting enzyme (“ICE”) is a cysteine protease whichcatalyzes the formation of interleukin-1β. Interleukin-1β is animmunoregulatory protein implicated in the following disorders:inflammation, diabetes, septic shock, rheumatoid arthritis, andAlzheimer's disease. ICE has also been linked to apoptotic cell death ofneurons, which is implicated in a variety of neurodegenerative disordersincluding Parkinson's disease, ischemia, and amyotrophic lateralsclerosis (ALS).

Cysteine proteases are also produced by various pathogens. The cysteineprotease clostripain is produced by Clostridium histolyticum. Otherproteases are produced by Trypanosoma cruzi, malaria parasitesPlasmodium falciparum and P. vinckei and Streptococcus. Hepatitis Aviral protease HAV C3 is a cysteine protease essential for processing ofpicornavirus structural proteins and enzymes.

Exemplary serine proteases implicated in degenerative disorders includethrombin, human leukocyte elastase, pancreatic elastase, chymase andcathepsin G. Specifically, thrombin is produced in the blood coagulationcascade, cleaves fibrinogen to form fibrin and activates Factor VIII;thrombin is implicated in thrombophlebitis, thrombosis and asthma. Humanleukocyte elastase is implicated in tissue degenerative disorders suchas rheumatoid arthritis, osteoarthritis, atherosclerosis, bronchitis,cystic fibrosis, and emphysema. Pancreatic elastase is implicated inpancreatitis. Chymase, an enzyme important in angiotensin synthesis, isimplicated in hypertension, myocardial infarction, and coronary heartdisease. Cathepsin G is implicated in abnormal connective tissuedegradation, particularly in the lung.

Given the link between cysteine and serine proteases and variousdebilitating disorders, compounds which inhibit these proteases would beuseful and would provide an advance in both research and clinicalmedicine. The present invention is directed to these, as well as other,important ends.

SUMMARY OF THE INVENTION

The present invention is directed to selected peptide-containingα-ketoamide inhibitors of cysteine and serine proteases represented bythe general formula I:

wherein:

Q has the formula G—B—(CHR⁴), where R⁴ is independently H or alkylhaving from 1 to 4 carbons;

v is 0, 1, or 2;

B is selected from the group consisting of C(═O), OC(═O), S(═O)_(m),CH₂, a bond, NR⁵C(═O), S(═O)_(m)—A—C(═O), and C(═O)—A—C(═O), where R⁵ isH or lower alkyl;

m is 0, 1, or 2;

A is lower alkylene or cycloalkylene, optionally substituted with one ormore halogen atoms, aryl, or heteroaryl groups;

M is a carbon atom;

G is selected from the group consisting of H, a blocking group, loweralkyl, lower alkenyl, aryl having from about 6 to about 14 carbons,heterocyclyl having from about 5 to about 14 ring atoms,heterocycloalkyl having from about 5 to about 14 ring atoms, arylalkylhaving from about 7 to about 15 carbons, heteroarylalkyl, andarylheteroalkyl wherein the aryl portion can be unfused or fused withthe heteroalkyl ring, said alkyl, aryl, heterocyclyl, heterocycloalkyl,arylalkyl, heteroarylalkyl, and arylheteroalkyl groups being optionallysubstituted with one or more J groups;

J is selected from the group consisting of halogen, CN, nitro, loweralkyl, cycloalkyl, heterocycloalkyl, heteroalkyl, halogenated alkyl,aryloxyalkyl, alkylthio, alkylsulfonyl, aryl, heteroaryl, arylalkyl,arylalkyloxy, arylsulfonyl, heteroarylsulfonyl, alkoxycarbonyl,alkoxyalkyl, acyl, alkoxy, hydroxy, carboxy, hydroxyalkyl, amino,alkylamino, and aminoalkyl, said amino group or said amino group of saidaminoalkyl or alkylamino group being optionally substituted with an acylgroup, an alkoxy group, or with 1 to 3 aryl, lower alkyl, cycloalkyl, oralkoxyalkyl groups; and said aryl, heteroaryl, heterocycloalkyl, andheteroalkyl groups being further optionally substituted by a J group;

each Aaa is independently an amino acid which optionally contains one ormore blocking groups;

n is 0, 1, 2, or 3;

R¹ and R² are independently selected from the group consisting of H,alkyl having from one to about 6 carbons, arylalkyl having from about 7to about 15 carbons, heteroalkyl in which the ring contains from about 5to about 14 ring atoms, heteroarylalkyl in which the heteroaryl ringcontains from about 5 to about 14 ring atoms, alkoxyalkyl, a side chainof a naturally occurring amino acid in the R or S configuration, and(CH₂)pNH-L, said alkyl, arylalkyl, heteroalkyl, heteroarylalkyl, andalkoxyalkyl groups being optionally substituted with one or more Jgroups;

p is 0, 1, 2, or 3;

L is selected from the group consisting of alkoxycarbonyl having from 2to about 7 carbons, arylalkoxycarbonyl in which the arylalkoxy groupcontains about 7 to about 15 carbons, and S(═O)₂R⁶;

R⁶ is selected from the group consisting of lower alkyl, and aryl havingfrom about 6 to about 14 carbons;

R³ is selected from the group consisting of H, alkyl having from one toabout 6 carbons, arylalkyl having from about 7 to about 15 carbons,heteroalkyl in which the ring contains from about 5 to about 14 ringatoms, heteroarylalkyl in which the heteroaryl ring contains from about5 to about 14 ring atoms, alkoxyalkyl, a side chain of a naturallyoccurring amino acid in the R or S configuration, (CH₂)_(p)NH—L,C(═O)R⁷, S(═O)₂R⁷, a blocking group, and when combined with the carbonatom to which RI is attached an alkylene group having from 2 to 5carbons, said alkylene group being optionally substituted with a groupselected from the group consisting of aryl, azide, CN, a protected aminogroup, and OS0₂-aryl, said alkyl, arylalkyl, heteroalkyl,heteroarylalkyl, and alkoxyalkyl groups being optionally substitutedwith one or more J groups;

R⁷ is selected from the group consisting of aryl having from about 6 toabout 14 carbons, heteroaryl having from about 5 to about 14 ring atoms,arylalkyl having from about 7 to about 15 carbons, alkyl having from 1to about 10 carbons, said aryl, heteroaryl, arylalkyl and alkyl groupsbeing optionally substituted with one or more J groups, heteroalkylhaving from 2 to about 7 carbons, alkoxy having from about 1 to about 10carbons, and amino optionally substituted with 1 or more alkyl groups;

q is 0 or 1;

Z is selected from the group consisting of C(═O)C(═O)NH—X—A¹—K and

X is a bond or —O—;

A¹ is the same as A;

K is selected from the group consisting of N(R¹⁰)Y,

and SO₂N(R⁸)(R¹⁰);

D is a fused aryl or heteroaryl group;

R¹¹ is selected from the group consising of alkoxy, aryloxy, and NHR¹²;

R¹² is selected from the group consisting of H, alkyl, aryl, andheteroaryl, said alkyl, aryl or heteroaryl groups being optionallysubstituted with one or more J groups;

Y is selected from the group consisting of SO₂R⁸, C(═O)NHR⁹, C(═S)NHR⁹,C(═NCN)R¹¹, C(═NC(═O)NHR¹⁰)R¹¹, and CO₂R⁸;

R⁸ is selected from the group consisting of alkyl, alkoxy, aryl, andheterocyclyl, said alkyl, alkoxy, aryl, or heterocyclyl groups beingoptionally substituted with one or more J groups;

R⁹ is selected from the group consisting of H, alkyl, aryl, andheteroaryl, said alkyl, aryl, or heteroaryl groups being optionallysubstituted with one or more J groups;

or an R⁹ alkyl group may be combined with an A¹ alkylene group to form aN-containing heterocyclic 5- or 6-membered ring;

R¹⁰ is selected from the group consisting of H and lower alkyl;

or in the moiety SO₂N(R⁸)R¹⁰I, R⁸ and R¹⁰ may be combined together withthe N atom to which they are attached to form a N-containingheterocyclic 5- or 6-membered ring;

or where A¹ is an alkylene group, and K is N(R¹⁰)Y wherein R¹⁰ is alkyl,said R¹⁰ alkyl group may be combined with said A¹ alkylene group to forma N-containing heterocyclic 5- or 6-membered ring; or a pharmaceuticallyacceptable salt thereof.

In some preferred embodiments of the compounds of Formula I, n and v areeach 0, q is 1, B is a bond, and G is H. In further preferredembodiments of the compounds of Formula I, R¹ is the sidechain of anaturally occurring amino acid. In still further preferred embodimentsof the compounds of Formula I, R³ is —S (═C)₂R⁸.

In some preferred embodiments of the compounds of Formula I, R² isbenzyl or alkoxyalkyl. In more preferred embodiments, X is a bond, and Yis SO₂R⁸. Preferably, A¹ is —CH₂—CH₂—, —CH₂—CH(CH₃)—, or —(CH₃)CH—CH₂—.

In further preferred embodiments of the compounds of Formula I, R¹ is aserine sidechain, which is optionally capped with a benzyl group.Preferably, the carbon to which the serine sidechain is attached,designated “M” in Formula I, is a carbon atom in the D configuration.

In preferred embodiments of the compounds of Formula I, R² is benzyl, R⁷is methyl, and R⁸ is substituted phenyl, unsubstituted phenyl,substituted heteroaryl, or unsubstituted heteroaryl. In particularlypreferred embodiments, R⁸ is aryl, aryl substituted with amino, arylsubstituted with heterocyclomethyl, heteroaryl, alkyl substituted withheteroaryl, or heteroaryl substituted with alkylthio, haloalkyl, alkyl,phenylsulfonyl, halogen, aminophenyl, amino, or dialkylaminoalkyl.

In further preferred embodiments of the compounds of Formula I, n, v andq are each 0, B is (C═O), and G is phenyl or lower alkyl, said phenyl orlower alkyl groups being optionally substituted with one or more Jgroups.

In more preferred embodiments of the invention, n and v are each 0, q is1, R¹ is the side chain of an amino acid in the D- or L-configuration,R³ is S(═O)₂R⁷, G is H, B is a bond, R² is benzyl or alkoxyalkyl, X is abond, and Y is SO₂R⁸.

In other preferred embodiments, A¹ is CH₂CH₂, CH₂CH(CH₃), or (CH₃)CHCH₂.In more preferred embodiments, R¹ is a serine side chain in theD-configuration in which the hydroxyl group is capped with benzyl, R² isbenzyl, R⁷ is methyl, and R⁸ is substituted or unsubstituted phenyl orsubstituted or unsubstituted heteroaryl.

More preferred are the substituents shown for R₁-R₄, B, G, Aaa, X, A¹,Y, n, q and v shown for the compounds in Tables 2, 3, 4 and 5.Especially preferred are the substituents shown for compounds 9, 13, 17,22, and 29-151.

Some especially preferred embodiments of the compounds of Formula I areshown in Tables 2, 3, 4 and 5, infra, with compounds 9, 13, 17, 22, and29-151 being particularly preferred.

Because the peptide-containing α-ketoamides of the invention inhibitcysteine proteases and serine proteases, they can be used in bothresearch and therapeutic settings.

In a research environment, preferred compounds having defined attributescan be used to screen for natural and synthetic compounds which evidencesimilar characteristics in inhibiting protease activity. The compoundscan also be used in the refinement of in vitro and in vivo models fordetermining the effects of inhibition of particular proteases onparticular cell types or biological conditions.

In a therapeutic setting, given the connection between cysteineproteases and certain defined disorders, and serine proteases andcertain defined disorders, compounds of the invention can be utilized toalleviate, mediate, reduce and/or prevent disorders which are associatedwith abnormal and/or aberrant activity of cysteine proteases and/orserine proteases.

In preferred embodiments, compositions are provided for inhibiting aserine protease or a cysteine protease comprising a compound of theinvention and a pharmaceutically acceptable carrier. In other preferredembodiments, methods are provided for inhibiting serine proteases orcysteine proteases comprising contacting a protease selected from thegroup consisting of serine proteases and cysteine proteases with aninhibitory amount of a compound of the invention.

Methodologies for making the present peptide-containing α-ketoamideinhibitors are also disclosed. Other useful methodologies will beapparent to those skilled in the art, once armed with the presentdisclosure. These and other features of the compounds of the subjectinvention are set forth in more detail below.

DETAILED DESCRIPTION

Disclosed herein are the selected peptide-containing α-ketoamides whichare represented by the following formula I:

wherein:

Q has the formula G—B—(CHR⁴)_(v) where R⁴ is independently H or alkylhaving from 1 to 4 carbons;

v is 0, 1, or 2;

B is selected from the group consisting of C(═O), OC(═O), S(═O)_(m),CH₂, a bond, NR⁵C(═O), S(═O)_(m)—A—C(═O), and C(═O)—A—C(═O), where R⁵ isH or lower alkyl;

m is 0, 1, or 2;

A is lower alkylene or cycloalkylene, optionally substituted with one ormore halogen atoms, aryl, or heteroaryl groups;

M is a carbon atom;

G is selected from the group consisting of H, a blocking group, loweralkyl, lower alkenyl, aryl having from about 6 to about 14 carbons,heterocyclyl having from about 5 to about 14 ring atoms,heterocycloalkyl having from about 5 to about 14 ring atoms, arylalkylhaving from about 7 to about 15 carbons, heteroarylalkyl, andarylheteroalkyl wherein the aryl portion can be unfused or fused withthe hetercalkyl ring, said alkyl, aryl, heterocyclyl, heterocycloalkyl,arylalkyl, heteroarylalkyl, and arylheteroalkyl groups being optionallysubstituted with one or more J groups;

J is selected from the group consisting of halogen, CN, nitro, loweralkyl, cycloalkyl, heterocycloalkyl, heteroalkyl, halogenated alkyl,aryloxyalkyl, alkylthio, alkylsulfonyl, aryl, heteroaryl, arylalkyl,arylalkyloxy, arylsulfonyl, heteroarylsulfonyl, alkoxycarbonyl,alkoxyalkyl, acyl, alkoxy, hydroxy, carboxy, hydroxyalkyl, amino,alkylamino, and aminoalkyl, said amino group or said amino group of saidaminoalkyl or alkylamino group being optionally substituted with an acylgroup, an alkoxy group, or with 1 to 3 aryl, lower alkyl, cycloalkyl, oralkoxyalkyl groups; and said aryl, heteroaryl, heterocycloalkyl, andheteroalkyl groups being further optionally substituted by a J group;

each Aaa is independently an amino acid which optionally contains one ormore blocking groups;

n is 0, 1, 2, or 3;

R¹ and R² are independently selected from the group consisting of H,alkyl having from one to about 6 carbons, arylalkyl having from about 7to about 15 carbons, heteroalkyl in which the ring contains from about 5to about 14 ring atoms, heteroarylalkyl in which the heteroaryl ringcontains from about 5 to about 14 ring atoms, alkoxyalkyl, a side chainof a naturally occurring amino acid in the R or S configuration, and(CH₂)_(p)NH—L, said alkyl, arylalkyl, heteroalkyl, heteroarylalkyl, andalkoxyalkyl groups being optionally substituted with one or more Jgroups;

p is 0, 1, 2, or 3;

L is selected from the group consisting of alkoxycarbonyl having from 2to about 7 carbons, arylalkoxycarbonyl in which the arylalkoxy groupcontains about 7 to about 15 carbons, and S(═C)₂R⁶;

R⁶ is selected from the group consisting of lower alkyl, and aryl havingfrom about 6 to about 14 carbons;

R³ is selected from the group consisting of H, alkyl having from one toabout 6 carbons, arylalkyl having from about 7 to about 15 carbons,heteroalkyl in which the ring contains from about 5 to about 14 ringatoms, heteroarylalkyl in which the heteroaryl ring contains from about5 to about 14 ring atoms, alkoxyalkyl, a side chain of a naturallyoccurring amino acid in the R or S configuration, (CH₂)_(p)NH—L,C(═O)R⁷, S(═)₂R⁷, a blocking group, and when combined with the carbonatom to which R¹ is attached an alkylene group having from 2 to 5carbons, said alkylene group being optionally substituted with a groupselected from the group consisting of aryl, azide, CN, a protected aminogroup, and OS0₂-aryl, said alkyl, arylalkyl, heteroalkyl,heteroarylalkyl, and alkoxyalkyl groups being optionally substitutedwith one or more J groups;

R⁷ is selected from the group consisting of aryl having from about 6 toabout 14 carbons, heteroaryl having from about 5 to about 14 ring atoms,arylalkyl having from about 7 to about 15 carbons, alkyl having from 1to about 10 carbons, said aryl, heteroaryl, arylalkyl and alkyl groupsbeing optionally substituted with one or more J groups, heteroalkylhaving from 2 to about 7 carbons, alkoxy having from about 1 to about 10carbons, and amino optionally substituted with 1 or more alkyl groups;

q is 0 or 1;

Z is selected from the group consisting of C(═O)C(═O)NH—X—A¹—K and

X is a bond or —O—;

A¹ is the same as A;

K is selected from the group consisting of N(R¹⁰)Y,

and SO₂N (R⁸)(R¹⁰);

D is a fused aryl or heteroaryl group;

R¹¹ is selected from the group consisting of alkoxy, aryloxy, and NHR¹²;

R¹² is selected from the group consisting of H. alkyl, aryl, andheteroaryl, said alkyl, aryl or heteroaryl groups being optionallysubstituted with one or more J groups;

Y is selected from the group consisting of SO₂R⁸, C(═O)NHR⁹, C(═S) NHR⁹,C(═NCN) R¹¹, C(═NC (═O)NHR¹⁰)R¹¹, and CO₂R⁸;

R⁸ is selected from the group consisting of alkyl, alkoxy, aryl, andheterocyclyl, said alkyl, alkoxy, aryl, or heterocyclyl groups beingoptionally substituted with one or more J groups;

R⁹ is selected from the group consisting of H, alkyl, aryl, andheteroaryl, said alkyl, aryl, or heteroaryl groups being optionallysubstituted with one or more J groups;

or an R⁹ alkyl group may be combined with an A¹ alkylene group to form aN-containing heterocyclic 5- or 6-membered ring;

R¹⁰ is selected from the group consisting of H and lower alkyl;

or in the moiety SO₂N(R⁸)R¹⁰, R⁸ and R¹⁰ may be combined together withthe N atom to which they are attached to form a N-containingheterocyclic 5- or 6-membered ring;

or where A¹ is an alkylene group, and K is N(R¹⁰)Y wherein R¹⁰ is alkyl,said R¹⁰ alkyl group may be combined with said A¹ alkylene group to forma N-containing hetrocyclic 5- or 6-membered ring; or a pharmaceuticallyacceptable salt thereof.

It is recognized that various stereoisomeric forms of the compounds ofFormula I may exist. Preferred compounds of the invention have any Aaagroups being a-amino acids in the L-configuration. However, racematesand individual enantiomers and mixtures thereof form part of the presentinvention.

The carbon atom designated as “M” in the compounds of Formula I canexist in either the D or the L configuration. In some preferredembodiments, M is a carbon atom having the “D” configuration.

As used herein, the term “alkyl” includes straight-chain, and branchedhydrocarbon groups such as, for example, methyl, ethyl, propyl,isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, pentyl,1-ethylpentyl, hexyl, and octyl groups. “Cycloalkyl” groups are cyclicalkyl groups, such as, for example, cyclopropyl, methylcyclopentyl, andcyclohexyl groups. Preferred alkyl groups have 1 to about 10 carbonatoms, most preferably “lower alkyl” of 1 to about 6 carbon atoms.“Alkylene” groups are alkyl groups having two points of attachment;i.e., non-terminal alkyl groups. “Lower alkylene” groups are branched orunbranched alkylene groups of 1 to about 6 carbon atoms such as, forexample, ethylene (—CH₂CH₂—), propylene, butylene, hexylene,1-methylethylene, 2-methylethylene, and 2-methylpropylene.“Cycloalkylene” groups are cyclic alkylene groups. “Acyl” groups arealkylcarbonyl groups. “Aryl” groups are aromatic cyclic compoundspreferably including but not limited to phenyl, tolyl, naphthyl,anthracyl, phenanthryl, and pyrenyl. Also included within the definitionof “aryl” are ring systems having two aromatic rings connected by abond, such as biphenyl. Preferred aryl groups include phenyl andnaphthyl.

The term “carbocyclic”, as used herein, refers to cyclic groups in whichthe ring portion is composed solely of carbon atoms. The term “halogen”refers to F, Cl, Br, and I atoms. The term “arylalkyl” denotes alkylgroups which bear aryl groups, for example, benzyl groups. As usedherein, “alkoxy” groups are alkyl groups linked through an oxygen atom.Examples of alkoxy groups include methoxy (—OCH₃) and ethoxy (—OCH₂CH₃)groups. In general, the term “oxy” when used as a suffix denotesattachment through an oxygen atom. Thus, alkoxycarbonyl groups arecarbonyl groups which contain an alkoxy substituent, i.e., groups ofgeneral formula —C(═O)—C—R, where R is alkyl. The term “alkoxyalkyl”denotes an alkoxy group attached to an alkyl group. The term “aryloxy”denotes an aryl group linked through an oxygen atom, and the term“arylalkyloxy” denotes an arylalkyl group linked through an oxygen atom.

The terms “heterocycle”, “heterocyclyl”, and “heterocyclic” refer tocyclic groups in which a ring portion includes at least one heteroatomsuch as O, N or S. Heterocyclic groups include “heteroaryl” as well as“heteroalkyl” groups. The term “heteroaryl” denotes aryl groups havingone or more hetero atoms (e.g., C, N, or S) contained within an aromaticring. Also included within the definition of “heteroaryl” are ringsystems having two aromatic rings connected by a bond, where at leastone of the rings contains a hetero atom. Preferred “heteroaryl” groupsinclude pyridyl, pyrimidyl, pyrrolyl, furyl, thienyl, imidazolyl,triazolyl, tetrazolyl, quinolyl, isoquinolyl, benzoimidazolyl,thiazolyl, bipyridyl, pyridylthiophenyl, pyrimidylthiophenyl,benzimidazolyl, isoxazolylthiophenyl, pyrazolylthiophenyl, phthalimido,and benzothiazolyl. The term “heterocycloalkyl” denotes a heterocycleattached through a lower alkyl group. The term “heteroarylalkyl” denotesa heteroaryl group attached through an alkyl group.

As used herein, the term “heteroalkyl” denotes a heterocyclic groupwhich contains at Least one saturated carbon atomn in a heterocyclicring. Examples of heteroalkyl groups include piperidine,dihydropyridine, and tetrahydroisoquinyl groups. The term“arylheteroalkyl” as used herein denotes a “heteroalkyl” group connectedthrough an aryl group. One preferred example of an arylheteroalkyl groupis dibenzo-γ-pyranyl.

As used herein, the term “amino acid” denotes a molecule containing bothan amino group and a carboxyl group. As used herein the term “L-aminoacid” denotes an α-amino acid having the L-configuration around theα-carbon, that is, a carboxylic acid of general formula CH(COOH)(NH₂)—(side chain), having the L-configuration. The term “D-amino acid”similarly denotes a carboxylic acid of general formulaCH(COOH)(NH₂)—(side chain), having the D-configuration around theα-carbon. Side chains of L-amino acids include naturally occurring andnon-naturally occurring moieties. Nonnaturally occurring (i.e.,unnatural). amino acid side chains are moieties that are used in placeof naturally occurring amino acid sidechains in, for example, amino acidanalogs. See, for example, Lehninger, Biochemistry, Second Edition,Worth Publishers, Inc, 1975, pages 73-75. One representative amino acidside chain is the lysyl side chain, —(CH₂)₄—NH₂. Other representativea-amino acid side chains are shown below in Table 1.

TABLE 1 CH₃— HS—CH₂— HO—CH₂— HO₂C—CH(NH₂)—CH₂S—S—CH₂— C₆H₅—CH₂— CH₃—CH₂—HO—C₆H₄—CH₂— CH₃S—CH₂—CH₂—

CH₃—CH₂—S—CH₂—CH₂— HO—CH₂—CH₂— CH₃—CH(OH)— HO₂C—CH₂—NHC(═O)—CH₂—

HO₂C—CH₂—CH₂— NH₂C(═O)—CH₂—CH₂—

(CH₃)₂—CH— (CH₃)₂—CH—CH₂— CH₃—CH₂—CH₂— H₂N—CH₂—CH₂—CH₂—

H₂N—C(═NH)—NH—CH₂—CH₂—CH₂— H₂N—C(═O)—NH—CH₂—CH₂—CH₂— CH₃—CH₂—CH(CH₃)—CH₃—CH₂—CH₂—CH₂— H₂N—CH₂—CH₂—CH₂—CH₂—

Functional groups present in the compounds of Formula I may containblocking groups. Blocking groups are known per se as chemical functionalgroups that can be selectively appended to functionalities, such ashydroxyl groups, amino groups, thio groups, and carboxyl groups.Protecting groups are blocking groups which can be readily removed fromfunctiorialities. These groups are present in a chemical compound torender such functionality inert to chemical reaction conditions to whichthe compound is exposed. Any of a variety of protecting groups may beemployed with the present invention. Examples of such protecting groupsare the benzyloxycarbonyl (Cbz; Z), toluenesulfonyl, t-butoxycarbonyl,methyl ester, and benzyl ether groups. Other preferred protecting groupsaccording to the invention may be found in Greene, T. W. and Wuts, P. G.M., “Protective Groups in Organic Synthesis” 2d. Ed., Wiley & Sons,1991, which is hereby incorporated by reference in its entirety.

Further blocking groups useful in the compounds of the present inventioninclude those that bear acyl, aroyl, alkyl, alkanesulfonyl,arylalkanesulfonyl, or arylsulfonyl substituents on their amino groups.Other useful blocking groups include alkyl ethers, e.g., the methylether of serine.

The disclosed compounds of the invention are useful for the inhibitionof cysteine proteases and serine proteases. As used herein, the terms“inhibit” and “inhibition” mean having an adverse effect on enzymaticactivity. An inhibitory amount is an amount of a compound of theinvention effective to inhibit a cysteine and/or serine protease.

Pharmaceutically acceptable salts of the cysteine and serine proteaseinhibitors also fall within the scope of the compounds as disclosedherein. The term “pharmaceutically acceptable salts” as used hereinmeans an inorganic acid addition salt such as hydrochloride, sulfate,and phosphate, or an organic acid addition salt such as acetate,maleate, fumarate, tartrate, and citrate. Examples of pharmaceuticallyacceptable metal salts are alkali metal salts such as sodium salt andpotassium salt, alkaline earth metal salts such as magnesium salt andcalcium salt, aluminum salt, and zinc salt. Examples of pharmaceuticallyacceptable organic amine addition salts are salts with morpholine andpiperidine. Examples of pharmaceutically acceptable amino acid additionsalts are salts with lysine, glycine, and phenylalanine.

Compounds provided herein can be formulated into pharmaceuticalcompositions by admixture with pharmaceutically acceptable nontoxicexcipients and carriers. As noted above, such compositions may beprepared for use in parenteral administration, particularly in the formof liquid solutions or suspensions; or oral administration, particularlyin the form of tablets or capsules; or intranasally, particularly in theform of powders, nasal drops, or aerosols; or dermally, via, forexample, transdermal patches; or prepared in other suitable fashions forthese and other forms of administration as will be apparent to thoseskilled in the art.

The composition may conveniently be administered in unit dosage form andmay be prepared by any of the methods well known in the pharmaceuticalart, for example, as described in Remington's Pharmaceutical Sciences(Mack Pub. Co., Easton, Pa., 1980). Formulations for parenteraladministration may contain as common excipients sterile water or saline,polyalkylene glycols such as polyethylene glycol, oils and vegetableorigin, hydrogenated naphthalenes and the like. In particular,biocompatible, biodegradable lactide polymer, lactide/glycolidecopolymer, or polyoxyethylene-polyoxypropylene copolymers may be usefulexcipients to control the release of the active compounds. Otherpotentially useful parenteral delivery systems for these activecompounds include ethylene-vinyl acetate copolymer particles, osmoticpumps, implantable infusion systems, cyclodextrins and liposomes.Formulations for inhalation administration contain as excipients, forexample, lactose, or may be aqueous solutions containing, for example,polyoxyethylene-9-lauryl ether, glycocholate and deoxycholate, or oilysolutions for administration in the form of nasal drops, or as a gel tobe applied intranasally. Formulations for parenteral administration mayalso include glycocholate for buccal administration, a salicylate forrectal administration, or citric acid for vaginal administration.Formulations for transdermal patches are preferably lipophilicemulsions.

The materials for this invention can be employed as the sole activeagent in a pharmaceutical or can be used in combination with otheractive ingredients which could facilitate inhibition of cysteine andserine proteases in diseases or disorders.

The concentrations of the compounds described herein in a therapeuticcomposition will vary depending upon a number of factors, including thedosage of the drug to be administered, the chemical characteristics(e.g., hydrophobicity) of the compounds employed, and the route ofadministration. In general terms, the compounds of this invention may beprovided in effective inhibitory amounts in an aqueous physiologicalbuffer solution containing about 0.1 to 10% w/v compound for parenteraladministration. Typical dose ranges are from about lug/kg to about 1g/kg of body weight per day; a preferred dose range is from about 0.01mg/kg to 100 mg/kg of body weight per day. Such formulations typicallyprovide inhibitory amounts of the compound of the invention. Thepreferred dosage of drug to be administered is likely, however, todepend on such variables as the type or extent of progression of thedisease or disorder, the overall health status of the particularpatient, the relative biological efficacy of the compound selected, andformulation of the compound excipient, and its route of administration.

As used herein, the term “contacting” means directly or indirectlycausing at least two moieties to come into physical association witheach other. Contacting thus includes physical acts such as placing themoieties together in a container, or administering moieties to apatient. Thus, for example administering a compound of the invention toa human patient evidencing a disease or disorder associated withabnormal and/or aberrant activity of such proteases falls within thescope of the definition of the term “contacting”.

The invention is further illustrated by way of the following exampleswhich are intended to elucidate the invention. These examples are notintended, nor are they to be construed, as limiting the scope of thedisclosure.

EXAMPLES

General Methods.

Thin layer chromatography was performed on silica gel plates (MK6F 60A,size 1×3 in, layer thickness 250 μm, Whatman Inc.). Preparative thinlayer chromatography was performed on silica gel plates (size 20×20 in,layer thickness 1000 micron, Analtech). Preparative columnchromatography was carried out using Merck silica gel, 40-63 μm, 230-400mesh. ¹H NMR spectra were recorded on a GE QE Plus instrument (300 MHZ)using tetramethylsilane as internal standard. Electrospray mass spectrawere recorded on a VG platform II instrument (Fisons Instruments).

Compounds of the invention were prepared following one of the GeneralMethods A, B, C or D.

Example 1

Preparation of Compound 9 by General Method A

Preparation of Compound 1

This compound and related hydroxy acids used in this study weresynthesized following a general procedure of Harbeson et al, J. Med.Chem. 1994, 37, 2918-2929, which is incorporated herein by reference inits entirety.

Preparation of Compound 2

To a cooled (−10° C.) solution of compound 1 (4.30 g, 0.015 mol) inanhydrous methanol (50 mL) was added slowly thionyl chloride (3.20 mL).After 0.5 hour, the cooling bath was removed, the mixture was stirredfor an additional 16 hours and concentrated to give a residue which ontrituration with ethyl acetate (30 mL) gave a white solid. The solid wasseparated by filtration and dried to give 3.50 g of compound 2 which wasused directly in the next step; MS m/e 210(M+H).

Preparation of Compound 3

The preparation of this compound is shown in General Method E.

Preparation of Compound 4

To a cooled (0° C.) solution of compound 1 (1.00 g, 0.0034 mol) inanhydrous DMF (20 mL) was added NMM (1.40 g, 0.014 mmol) followed by1-HOBt (0.54 g, 0.0040 mmol) and BOP (1.80 g, 0.0040 mmol). The mixturewas stirred for 15 minutes and to it was added compound 3 (0.75g, 0.0032mmol). The cooling bath was removed and the mixture was stirred for 4hours, poured into ice-water (200 mL), and extracted into ethyl acetate(3×100 mL). The organic layer was washed with 2% citric acid solution(2×50 mL), 2% sodium bicarbonate solution (2×50 mL), brine (1×50 mL),and it was dried over anhydrous sodium sulfate. Solvent evaporationunder reduced pressure gave a crude solid which was washed several timeswith n-pentane to produce 1.30 g of compound 4.

Compound 4: white solid (diastereomeric mixture); ¹H-NMR (DMSO-d₆) δ7.90 and 7.65 (2 sets of t, 1H), 7.75 (d, 2H), 7.55 (q, 2H), 7.15 (m,6H), 6.55 and 5.80 (2 sets of d, 1H), 3.90 (m, 2H), 3.30 (d, 1H), 3.10(m, 2H), 2.75 (m, 2H), 2.50 (m, 3H), 1.20 (s, 9H). MS m/e 478(M+H),500(M+Na).

Preparation of Compound 5

To a solution of compound 4 (0.40 g, 0.84 mmol) in 1,4-dioxane (15 mL)was added 4 N HCl in dioxane (15 mL). The reaction mixture was stirredat room temperature for 2 hours, then concentrated at reduced pressureto give a residue which was washed several times with ethyl acetate anddried under vacuum to give 0.30 g of compound 5; ¹H-NMR (DMSO-d₆) showedcomplete absence of tBoc peak at δ 1.20 ppm; MS m/e 378 (M+H). Thismaterial was used directly in the next step.

Preparation of Compound 7

To a mixture of D-Ser(Bn) (compound 6, 1.00g, 5 mmol) and 1 N NaOH (10mL, 10 mmol) at 0° C. was slowly added methanesulfonyl chloride (0.80 g,7.69 mmol). After 0.5 hour, the cooling bath was removed, the mixturewas stirred overnight and acidified (pH˜2-3) with 2 N HCl. The aqueouslayer was extracted into ethyl acetate (3×50 mL). The combined organiclayer was washed with water (1×20 mL) and brine (1×20 mL), and driedover MgSO₄. Solvent evaporation gave a residue which was redissolved inmethylene chloride (10 mL); addition of hexanes produced a white solidwhich was filtered and dried to give 1.02 g of compound 7.

Preparation of Compound 8

This compound was prepared by coupling compound 7 and compound 5, usingNMM/HOBt/BOP as coupling agents, following the procedure described abovefor the preparation of compound 4. In some of the related examples,EDCI/HOBt were used as coupling agents.

Preparation of Compound 9

To a cooled (0° C.) solution of compound 8 (0.31 g, 0.49 mmol) inanhydrous methylene chloride (10 mL) was added Dess-Martin periodinanereagent (0.425 g, 1.00 mmol). The cooling bath was removed and themixture was stirred for an additional 1 hour. The solution was thendiluted with methylene chloride (10 mL), and washed with 10% sodiumthiosulfate solution (5×5 ml), saturated sodium bicarbonate solution(2×5 mL), and brine (1×5 mL), and dried over anhydrous sodium sulfate.Solvent removal under reduced pressure gave a residue which was washedwith n-pentane (10 mL) and dried under vacuum to produce 0.178 g ofcompound 9; ¹H-NMR spectrum revealed a minor amount of epimerization hadtaken place.

Compound 9: white solid; ¹H-NMR (DMSO-d₆) δ 8.75 (t, 1H), 8.60 and 8.50(2 doublets, 1H), 7.75 (d, 2H), 7.65-7.00 (a series of m, 15H), 5.25(broad m, 1H), 4.45 and 4.235 (2 singlets, 2H), 4.15 (m, 1H), 3.35-2.60(a series of m, 8H), 3.35 and 3.25 (2 singlets, 3H) MS m/e 631(M+H),653(M+Na).

Example 2

Preparation of Compound 13 by General Method B

In General Method B, compound 4, prepared as described above, wasoxidized by Dess-Martin periodinane reagent to generate compound 10which on tBoc-deprotection (2 N HCl in dioxane) produced the amine-salt,compound 11. Coupling (NMM/HOBt/BOP) of compound 11 withN-phenylsulfonyl-(L)-Pro (compound 12) yielded compound 13. Purificationwas achieved by passing a solution of the crude material in methylenechloride through Sep-Pak® Vac 6 cc (1 g) silica cartidge (WatersCorporation, Milford, Mass.), eluting with methylene chloride, followedby various combinations of methylene chloride and ethyl acetate.Harbeson et al. (J. Med. Chem. 1994, 37, 2918-2929) reported that silicagel chromatography of a ketoamide epimerizes the chiral center at P₁.

Compound 13: white solid; ¹H-NMR (CDCl₃) δ 7.90-7.00 (a series of m,18H), 5.40 and 5.30 (2 multiplets, 1H), 4.10 (m, 1H), 3.50-3.00 (m, 8H),1.90-1.40 (m, 4H). MS m/e 613 (M+H), 635 (M+Na)

Example 3

Preparation of Compound 17 by General Method C

In General Method C, compound 2 was coupled (NMM/HOBt/BOP) withL-Cbz-Leu to give compound 14 which was hydrolyzed (aq. NaOH) tocompound 15. Coupling (NMM/HOBt/BOP) of compound 15 with compound 3 gavecompound 16 which underwent Dess-Martin oxidation to generate compound17.

Compound 17: white solid; ¹H-NMR (CDCl₃) δ 7.85 (d, 2H), 7.60-7.00 (aseries of m, 15H), 6.60 (d, 1H), 5.40 (m, 1H), 5.20 (q, 1H), 5.10 (s,2H), 4.10 (broad, 1H), 3.50-3.00 (a series of m, 6H), 1.65-1.30 (m, 3H),0.90 (d, 6H). MS m/e 623 (M+H), 645 (M+Na).

Example 4

Preparation of Compound 22 by General Method D

In General Method D, compound 7 was coupled (NMM/HOBt/BOP) with compound2 to generate compound 18 which underwent Dess-Martin oxidation togenerate compound 19. Hydrolysis (LiOH, MeOH—H₂O) of compound 19 gavecompound 20 which was coupled (NMM/HOBt/BOP) with compound 21 to givecompound 22. Compound 22 was purified by silica gel chromatography.

Compound 22: white solid; MS m/e 646(M+H), 668(M+Na).

Preparation of Intermediates

Example 5

The preparation of a representative example of an amine (compound 3),containing a terminal sulfonamide moiety, is shown in General Method E.

Preparation of Compound 24

To a solution of 1,2-ethylenediamine (compound 23, 10.80 g, 12.00 mL,0.18 mol) in THF (30 mL) was added slowly BOC-ON (22.10 g, 0.09 mol) inTHF (70 mL) over a period of 4 hours. The reaction mixture was stirredovernight, concentrated on a rotavapor, and taken up into water (150mL). The aqueous layer was acidified (pH˜5-6) with solid citric acidmonohydrate, washed with ether (3×50 mL) and then treated (at 0° C.)with 6 N NaOH solution to make it basic (pH˜12-13). The basic solutionwas extracted into ethyl acetate (3×100 mL), and the combined ethylacetate layer was dried (MgSO₄) and concentrated to generate 7.23 g ofmonoprotected diamine, compound 24. Compound 24: viscous liquid; ¹H-NMR(CDCl₃) δ 5.00 (broad, 1H), 3.20 (broad q, 2H), 2.80 (t, 2H), 1.45 (s,9H), 1.25 (broad, 2H).

Preparation of Compound 25

A cooled (0-5° C.) solution of compound 24 (0.321 g, 0.002 mol) inmethylene chloride (5 mL) was treated sequentially with triethylamine(0.243 g, 0.33 mL, 0.0024 mol) and benzenesulfonyl chloride (0.423 g,0.30 mL, 0.0024 mol). The ice-bath was removed and the mixture wasstirred for an additional 0.5 hour, washed successively with water (2×5mL), cold (0-5° C.) 0.5 N HCl (1×5 mL), 2% NaHCO₃ solution (1×5 mL), andbrine (1×5 mL). The solution was dried (MgSO₄) and the solvent wasevaporated to give a residue which was washed several times withn-pentane. A total of 0.60 g of the sulfonamide derivative, compound 25,was obtained.

Compound 25: white solid, mp 92-95° C.; Rf (TLC, 5% methanol inmethylene chloride) 0.55; ¹H-NMR (CDCl₃) δ 7.85 (d, 2H), 7.55 (m, 3H),5.30 (broad d, 1H), 4.85 (broad, 1H), 3.25 (broad q, 2H), 3.10 (broad q,2H), 1.40 (s, 9H).

Preparation of Compound 3

A solution of compound 25 (0.560 g, 0.0019 mol) in 1,4-dioxane (4 mL)was treated with 4 N HCl in dioxane (4 mL). The mixture was stirred atroom temperature for 1 hour and concentrated at the rotavapor. Theresidue was washed several times with ethyl acetate and dried undervacuum to give 0.40 g of compound 3.

Compound 3: white solid, mp 178-180° C.; ¹H-NMR (DMSO-d₆)δ 8.20-8.00(broad t, 4H), 7.80 (d, 2H), 7.60 (m, 3H), 2.95 (broad q, 2H), 2.80(broad, 2H).

Example 6

Preparation of Compound 28

The preparation of a representative example of an intermediate amine(compound 28) containing a terminal biaryl sulfonamide moiety is shownin General Method F.

A mixture of compound 26 (prepared from compound 24 and5-bromothiophene-2-sulfonyl chloride, following the same generalprocedure as described above for the preparation of compound 25, 0.50 g,1 eqv), dimethoxyethane (10 mL), 2 M Na₂CO₃ (5 eqv), phenylboronic acid(1.40 eqv) and Pd(PPh₃)₄ (0.04 eqv) was heated at 135 IC for 2.5 hours.The reaction mixture was concentrated at the rotavapor, and the residuewas taken up into water (20 mL). The aqueous layer was acidified withcitric acid and extracted into methylene chloride (3×20 mL). Thecombined organic layer was washed with water (1×10 mL) and brine (1×10mL). It was dried (MgSO₄) and concentrated to a small volume.Trituration of the residue with hexanes gave a solid which was separatedby filtration and dried under vacuum to produce 0.37 g of compound 27;¹H-NMR (CDCl₃) δ 7.60-7.20 (a series of m, 7H), 5.35 (broad, 1H), 4.85(broad, 1H), 3.30 (m, 2H), 3.20 (m, 2H), 1.40 (s, 9H). For a generaldescription of this reaction procedure, see Miyaura et al., Chem. Rev.1995, 95, 2457-2483.

Compound 27 was converted to compound 28 following the proceduredescribed for the preparation of compound 3.

Example 7

Preparation of Taurine Sulfonamide Intermediate

The preparation of a representative taurine sulfonamide intermediate isshown in General Method G.

The phthalimide of taurine, prepared by a known procedure (R.Winterbottom et al., J. Amer. Chem. Soc., 1947, 69, 1393-1401) wasconverted to its sulfonyl chloride with phosphorous pentachloride inrefluxing benzene. This was allowed to react with aniline in thepresence of pyridine to form the corresponding sulfonamide. Thephthalimide protecting group was then removed by refluxing withhydrazine and the resulting taurine sulfonamide was isolated as itshydrochloride.

Example 8

Syntheses of compounds 29 through 50 in Tables 2 and 3 were carried outusing the designated general methods, as described, and the appropriatestarting materials.

Example 9

Inhibition of Cysteine Protease Activity

To evaluate inhibitory activity, stock solutions (40 times concentrated)of each compound to be tested were prepared in 100% anhydrous DMSO and 5μl of each inhibitor preparation were aliquoted into each of three wellsof a 96-well plate. Recombinant human calpain I, prepared by the methodof Meyer et al. (Biochem. J. 1996, 314: 511-519; incorporated herein byreference in its entirety), was diluted into assay buffer (i.e., 50mMTris, 5OmM NaCl, lmM EDTA, 1 mM EGTA, and 5mM 8-mercaptoethanol, pH 7.5,including 0.2 mM Succ-Leu-Tyr-MNA), and 175 μl was aliquoted into thesame wells containing the independent inhibitor stocks as well as topositive control wells containing 5 μl DMSO, but no compound. To startthe reaction, 20 μl of 50 mM CaCl₂ in assay buffer was added to allwells of the plate, excepting three, which were used as backgroundsignal baseline controls. Substrate hydrolysis was monitored every 5minutes for a total of 30 minutes. Substrate hydrolysis in the absenceof inhibitor was linear for up to 15 minutes.

Inhibition of calpain I activity was calculated as the percent decreasein the rate of substrate hydrolysis in the presence of inhibitorrelative to the rate in its absence. Comparison between the inhibitedand control rates was made within the linear range for substratehydrolysis. The IC₅₀s of inhibitors (concentration yielding 50%inhibition) were determined from the percent decrease in rates ofsubstrate hydrolysis in the presence of five to seven differentconcentrations of the test compound. The results were plotted as percentinhibition versus log inhibitor concentration, and the IC₅₀ wascalculated by fitting the data to the four-parameter logistic equationshown below using the program GraphPad Prism (GraphPad Software, Inc.,San Diego, Calif.).

y=d+[(a−d)/(1+(x/c)^(b))]

The parameters a, b, c, and d are defined as follows: a is % inhibitionin the absence of inhibitor, b is the slope, c is the IC₅₀, and d is theE inhibition at an infinite concentration of inhibitor.

Results are presented in Tables 2, 3, 4 and 5 below.

TABLE 2 Inhibitory Activity of Linear α-Ketoamides

Cmp. Calpain Prep. MS No. W R₂ R IC₅₀ nM Method (M + 1)+  9 Ms-D- Bn Ph16 A 631 Ser(Bn) 13 PhSO₂-L- Bn Ph (78%) B 613 Pro ** 17 Cbz-L-Leu Bn Ph11 C 623 22 Ms-D- Ser(Bn) Bn

42 D 646 29 Ms-D- Ser(Bn) Bn

14 A 714 30 PhSO₂-L- Bn Ph (97%) B 663 Phe ** 31 Ms-D- Ser(Bn) Bn

29 A 632 32 Ms-D- Ser(Bn) Bn

10 A 704 33 Ms-D- Ser(Bn) Bn

17 A 761 34 Ms-D- Ser(Bn) Bn

25 A 786 (M + 2)⁺ 35 Ms-D- Bn CH₃ 91 A 569 Ser(Bn)  36* Ms-D- Ser(Bn)CH₂OMe

14 A 668 37 Ms-D- Ser(Bn) Bn

18 C 777  38* Ms-D- CH₂OMe Ph (100%) A 585 Ser(Bn) * 39 Ms-D- Ser(Bn) Bn

22 C 715, 717 ⁷⁹Br, ⁸¹Br 40 Ms-D- Ser(Bn) Bn

63 D 722 41 Ms-D- Ser(Bn) Bn

(88%) ** D 699 42 Ms-D- Ser(Bn) Bn

14 C 712(M)⁺ 43 Ms-D- Ser(Bn) Bn

11 C 718(M)⁺ 44 Ms-D- Ser(Bn) Bn

31 D 729 (M + 2)⁺ 45 Ms-D- Ser(Bn) Bn

64 D 688 46 Ms-D- Ser(Bn) Bn

144 D 660 *2:1 ratio of diastereomers. **Percent inhibition @ 10 μM

TABLE 3 Inhibitory Activity of Branched-Chain α-Ketoamides

Ex. Calpain IC₅₀ Prep. No. D nM Method MS (M + 1) 47

26 A 645 48

43 A 645 49

32 A 646 (M + 2) 50 CH₂CH₂N(CH₃)SO₂Ph (100%) C 645 **

Compounds listed in Table 4 were prepared by the general methods A-Gdescribed above.

TABLE 4 Inhibitory Activity of α-Ketoamides

Ex. Calpain MS No. W R IC₅₀ nM (M + 1) 51 Ms-D-SerCH₂CH₂NHSO₂-(3-(2-NH₂-thiazol-4-yl)Ph) 29 729 (Bn) 52 Ms-D-SerCH₂CH₂NHSO₂-(5-(3-formylphenyl)thiophene-  5 741 (Bn) 2-yl) 53 Ms-D-SerCH₂CH₂NHC(═N—CN)OPh 15 635 (Bn) 54 Ms-D-Ser CH₂CH₂NHSO₂(5-(3- 12 770(Bn) (Me₂NCH₂)phenyl)thiophene-2-yl) 55 Ms-D-Ser 3-Boc-NH-cyclohexane 42645 (Bn) 56 Ms-D-Ser CH₂CH₂NHSO₂(5-(3- 18 812 (Bn)(morpholinoCH₂)phenyl)thiophene-2-yl) 57 Ms-D-SerCH₂CH₂NHSO₂(4-(MorpholinoCH₂)Ph) 18 730 (Bn) 58 Ms-D-SerCH₂CH₂NHSO₂(5-(3-(N-Me-piperazinyl- 21 825 (Bn)CH₂)phenyl)thiophene-2-yl) 59 Ms-D-Ser CH₂CH₂NHSO₂(5-(3- 35 765 (Bn)(HOCH₂)phenyl)thiophene-2-yl)(2 diast) (M + Na) 60 Ms-D-SerCH₂CH₂SO₂NHPh 47 631 (Bn) 61 Ms-D-Ser CH₂CH₂SO₂NH(4-CF₃Ph) 32 699 (Bn)62 Ms-D-Ser (CH₂)₃SO₂NHPh 18 645 (Bn) 63 Ms-D-Ser (CH₂)₃SO₂NH(4-CF₃Ph)23 713 (Bn) 64 Ms-D-Ser 6-ketopiperidin-3-yl  (33)* 545 (Bn) 65 Ms-D-SerCH₂CH₂N(Me)SO₂-(5-(3- 19 755 (Bn) formylphenyl)thiophene-2-yl) 66Ms-D-Thr CH₂CH₂NHSO₂(5-pyrid-2-ylthiophene-2-yl) 12 728 (Bn) 67 Ms-D-Ser—N(Me)SO₂-(5-isoxazol-3-yl-thiophene-2-yl) 21 718 (Bn) 68 Ms-(D,L)-CH₂CH₂NHSO₂(5-pyrid-2-ylthiophene-2-yl) 21 670 Phenylgly 69 Ms-(D,L)-CH₂CH₂NHSO₂Ph 80 587 Phenylgly 70 Ms-D-Thr CH₂CH₂NHSO₂(5-(3- 23 826 (Bn)(morpholinoCH₂)phenyl)thiophene-2-yl) (Mixture of diastereomers) 71Ms-D-Phe CH₂CH₂NHSO₂(5-pyrid-2-ylthiophene-2-yl) 18 684 72 Ms-D-SerCH₂CH₂NHSO₂(5-(3-Fluorophenyl)thiophene-2- 18 731 (Bn) yl) (Mixture ofdiastereomers) 73 Ms-D-Ser (CH₂)₃SO₂NHOCH₃ 87 597 (Bn) (M − 1) 74Ms-D-Ser CH₂CH₂NHSO₂-(5-(3-Nitrophenyl)thiophene-2- 15 758 (Bn) yl)(Mixture of diastereomers) 75 Ms-D-SerCH₂CH₂NHSO₂-(5-(3-Methylphenyl)thiophene- 36 727 (Bn) 2-yl) (Mixture ofdiastereomers) 76 Ms-D-Ser CH₂CH₂NHSO₂(5-(3-(AcNH)phenyl)thiophene- 11792 (Bn) 2-yl) (M + Na) (Mixture of diastereomers) 77 Ms-D-SerCH₂CH₂NHSO₂(5-(3- 10 777 (Bn) (CH₃CO)phenyl)thiophene-2-yl) (M + Na)(Mixture of diastereomers) 78 Ms-D-Ser 1-(4- 48 770 (Bn)(Morpholinomethyl)benzenesulfonyl)pip eridin-4-yl) (Mixture ofdiastereomers) 79 Ms-D-Ser CH₂CH₂NHSO₂((4- 11 847 (Bn)(CH₃COPh)piperazin-1-yl)CH₂Ph) (Mixture of diastereomers) 80 Ms-D-Ser(CH₂)₃SO₂NH-morpholin-4-yl 169  652 (Bn) (M − 1) 81 Ms-D-Ser(CH₂)₃SO₂-morpholin-4-yl 124  639 (Bn) 82 Ms-D-Ser CH₂CH₂NHSO₂-(5-(4- 13765 (Bn) Methoxyphenyl)thiophene-2-yl) (M + Na) (Mixture ofdiastereomers) 83 Ms-D-Ser CH₂CH₂CH₂-Saccharin 48 657 (Bn) 84 Ms-D-SerCH₂CH₂NHSO₂((4-(PhCH₂)piperazin-1- 23 819 (Bn) yl)CH₂Ph) 85 Ms-D-SerCH₂CH₂NHSO₂((4-(CH₃CO)piperazin-1- 14 771 (Bn) yl)CH₂Ph) 86 Ms-D-SerCH₂CH₂NHSO₂-(5-Me₂N-naphth-1-yl) 49 724 (Bn) 87 Ms-D-SerCH₂CH₂NHSO₂-benzothiophene-2-yl 23 687 (Bn) 88 Cbz-Leu-CH₂CH₂NHSO₂(5-pyrid-2-ylthiophene- 33 819 Leu 2-yl) 89 Ms-D-SerCH₂CH₂NHSO₂((4-Pyrid-2-yl)piperazin- 21 806 (Bn) 1-yl)CH₂Ph) (Mixture ofdiastereomers) 90 Ms-D-Ser CH₂CH₂NHSO₂-(5-(4- 17 741 (Bn)formylphenyl)thiophene-2-yl) 91 Ms-D-Ser CH₂CH₂NHSO₂(4-(2- 19 758 (Bn)(MeOCH₂)PyrrolidinylCH₂)Ph) 92 Ms-D-Ser(CH₂)₅NHSO₂(5-pyrid-2-ylthiophene-2- 12 756 (Bn) yl) (Mixture ofdiastereomers) 93 Ms-D-Ser CH₂CH₂NHSO₂(5-(2- 40 812 (Bn)(morpholinoCH₂)phenyl)thiophene-2-yl) 94 Ms-D-Ser CH₂CH₂NHSO₂(5-(4- 22812 (Bn) (morpholinoCH₂)phenyl)thiophene-2-yl) 95 Ms-D-SerCH₂CH₂NHSO₂(5-(3- 30 810 (Bn) (piperidinylCH₂)phenyl)thiophene-2-yl) 96Ms-D-Ser CH₂CH₂NHSO₂(2-acetamido-4- 23 709 (Bn) methylthiazol-5-yl) 97Ms-D-Ser CH₂CH₂NHSO₂(1- 32 671 (Bn) phenylsulfonylpiperidin-4-yl) 98Ms-D-Ser CH₂CH₂NHSO₂-(5-(2- 24 741 (Bn) formylphenyl)thiophene-2-yl) 99Ms-D-Ser CH₂CH₂NHSO₂((CH₃O)CH₃NCH₂Ph) 21 704 (Bn) (Mixture ofdiastereomers) 100  Ms-D-Ser CH₂CH₂NHSO₂(4-ethylpiperazin-1- 21 756 (Bn)yl)CH₂Ph) (Mixture of diastereomers) 101  Ms-D-Ser CH₂CH₂NHSO₂(5-(3- 22798 (Bn) (Et₂NCH₂)phenyl)thiophene-2-yl) 102  Ms-D-Ser CH₂CH₂NHSO₂(5-(3-36 838 (Bn) (Cyclohexyl(Me)NCH₂)phenyl)thiophen e-2-yl) 103  Ms-D-SerCH₂CH₂NHSO₂(5-(3- 24 796 (Bn) (pyrrolidinylCH₂)phenyl)thiophene-2-yl)104  Ms-D-Ser CH₂CH₂NHSO₂(5-(3- 10 738 (Bn) cyanophenyl)thiophene-2-yl)105  Ms-D-Ser CH₂CH₂NHSO₂(4-(4- 14 816 (Bn) acetamidophenoxy)CH₂Ph) (M +Na) 106  Ms-D-Ser CH₂CH₂NHSO₂(5-(3- 44 782 (Bn)(azetidinylCH₂)phenyl)thiophene-2-yl) 107  Ms-D-Ser1-(5-pyridin-2-ylthiophene-2-yl- 23 754 (Bn) SO₂)Piperidin-4-yl)(Mixture of diastereomers) 108  Ms-D-SerCONHCH₂CH₂NHSO₂(5-(3-(N-ethyl-N- 10 784 (Bn)methylaminomethyl)phenyl)thiophene- 2-yl) 109  Ms-D-SerCH₂CH₂NHSO₂(5-(3-(bis(2- 22 858 (Bn)methoxyethyl)aminomethyl)phenyl)thiophene-2- yl) 110  Ms-D-SerCH₂CH₂NHSO₂(5-(3-cyanophenyl)thiophene-2- 11 738 (Bn) yl) (Mixture ofdiastereomers) 111  Ms-D-Ser CH₂CH₂NHSO₂(4-(3-pyrrolin-1-yl)CH₂Ph) 73712 (Bn) (Mixture of diastereomers) 112  Ms-D-SerCH₂CH₂NHSO₂((4-(CH₃SO₂)piperazin-1- 37 807 (Bn) yl)CH₂Ph) 113  Ms-D-SerCH₂CH₂NHSO₂((4-pyrimid-2-yl)piperazin-1- 24 807 (Bn) yl)CH₂Ph 114 Ms-D-Ser CH₂CH₂NHSO₂(5-(3- 33 828 (Bn)(thiomorpholinoCH₂)phenyl)thiophene-2-yl) 115  Ms-D-SerCH₂CH₂NHSO₂(5-(3-(4- 16 824 (Bn)ketopiperidinylCH₂)phenyl)thiophene-2-yl) 116  Ms-L-Ser CH₂CH₂NHSO₂Ph100  631 (Bn) *Percent inhibition @ 0.1 μM Ms = methylsulfonyl

TABLE 5 Inhibitory Activity of Achiral P₂ Mimetic α-Ketoamides

Ex. Calpain Synthesis MS No. Q R IC₅₀ nM Method (M + 1)⁺ 117 Benzoyl

800 A 563 118 2,6-Dichlorobenzoyl

 36 A 631, 633 119 2,6-Dichloro-3- methylbenzoyl

 61 A 645 120 2,6-Difluorobenzoyl

 20 A 599 121 2,4,6-Trifluorobenzoyl

 85 A 618 122 2,3,4,5,6-Pentafluoro- benzoyl

 28 A 653 123 3,4-Methylenedioxy- benzoyl

>1000  A 607 124 2,5-Dichlorobenzoyl

 68 A 631, 633 125 2-Chloro-5- methoxybenzoyl

 65 A 627 126 3,5-bis(trifluoro- methyl)benzoyl

600 A 699 127 2,6-Dimethylbenzoyl

178 A 591 128 2,6-Dichloronicotinoyl

 80 A 634, 636 129 2,6-Dichlorobenzoyl

 21 A 658, 660 130 2,6-Dichlorobenzoyl

 20 A 687, 689 131 2,6-Dichlorobenzoyl

 29 A 729, 731 132 2,6-Dichlorobenzoyl

 83 A 723, 725 133 2,6-Dichlorobenzoyl

 11 A 655, 657 134 2,6-Dichlorobenzoyl

100 A 688, 690 135 2,6-Difluorobenzoyl

 22 A 645 (M + Na) 136 2,6-Difluorobenzoyl

 62 A 611 (M + Na) 137 2,6-Diethylbenzoyl

145 A 631 (M + Na) 138 2,6-Dimethoxybenzoyl

4000  A 613 139 2-Isopropylbenzoyl

168 A 595 140 2-Chloro-6- fluorobenzoyl

 58 A 605 141 2-Fluoro-6-trifluoro- methylbenzoyl

 58 A 639 142 2,3,4,5,6-Pentafluoro- benzoyl

 32 A 643 143 2-Methylpropanoyl

1500  A 529 144 3-Methylbutanoyl

590 A 543 145 4-Methylpentanoyl

 29 A 557 146 3- Cyclopentylpropanoyl

1000  A 583 147 E-3-Hexenoyl

1000  A 555 148 4-Phenylpentanoyl

 87 A 641 (M + Na) 149 4-Phenylbutanoyl

1500  A 627 (M + Na) 150 4-Methylpentanoyl

 15 A 603 (M + Na) 151 3- Cyclopentylpropanoyl

420 A 607

As those skilled in the art will appreciate, numerous changes andmodifications may be made to the preferred embodiments of the inventionwithout departing from the spirit of the invention. It is intended thatall such variations fall within the scope of the invention.

It is intended that each of the patents, applications, and printedpublications mentioned in this specification be hereby incorporated byreference in their entirety.

What is claimed is:
 1. A compound having the Formula:

wherein D is CH₂CH₂N(CH₃)SO₂Ph or has one of the formulas:


2. A compound having the Formula:

wherein: R₂ is independently selected from the group consisting of Bnand CH₂OMe; and W and R are independently selected from the groupconsisting of Ms-D-Ser(Bn), Ph, PhSO₂-L-Pro, Cbz-L-Leu,


3. A compound having the formula:

wherein W and R are independently selected from the group consisting ofMs-D-Ser (Bn), CH₂CH₂NHSO₂—(3-(2-NH₂-thiazol-4-yl)Ph),CH₂CH₂NHSO₂-(5-(3-formylphenyl)thiophene-2-yl),CH₂CH₂NHSO₂(5-(3-(Me₂NCH₂)phenyl)thiophene-2-yl),CH₂CH₂NHSO₂(5-(3-(morpholinoCH₂)phenyl)thiophene-2-yl),CH₂CH₂NHSO₂(4-(MorpholinoCH₂)Ph), CH₂CH₂NHSO₂(5-(3-(N-Me-piperazinyl-CH₂)phenyl)thiophene-2-yl),CH₂CH₂NHSO₂(5-(3-(HOCH₂)phenyl)thiophene-2-yl), CH₂CH₂SO₂NHPh,CH₂CH₂SO₂NH(4-CF₃Ph), (CH₂)₃SO₂NHPh, (CH₂)₃SO₂NH(4—CF₃Ph),CH₂CH₂N(Me)SO₂-(5-(3-formylphenyl)thiophene-2-yl), Ms-D-Thr (Bn),CH₂CH₂NHS0₂(5-pyrid-2-ylthiophene-2-yl),—N(Me)SO₂-(5-isoxazol-3-yl-thiophene-2-yl), Ms-(D,L)-Phenylgly,CH₂CH₂NHSO₂(5-pyrid-2-ylthiophene-2-yl), CH₂CH₂NHSO₂Ph, Ms-D-Thr (Bn),CH₂CH₂NHSO₂(5-(3-(morpholinoCH₂)phenyl)thiophene-2-yl), Ms-D-Phe,CH₂CH₂NHSO₂(5-pyrid-2-ylthiophene-2-yl),CH₂CH₂NHSO₂(5-(3-Fluorophenyl)thiophene-2-yl),CH₂CH₂NHSO₂-(5-(3-Nitrophenyl)thiophene-2-yl),CH₂CH₂NHSO₂-(5-(3-Methylphenyl)thiophene-2-yl), CH₂CH₂NHSO₂(5-(3-(AcNH)phenyl)thiophene-2-yl),CH₂CH₂NHSO₂(5-(3-(CH₃CO)phenyl)thiophene-2-yl),CH₂CH₂NHSO₂-(5-(4-Methoxyphenyl)thiophene-2-yl), Cbz-Leu-Leu,CH₂CH₂NHSO₂ (5-pyrid-2-ylthiophene-2-yl),CH₂CH₂NHSO₂-(5-(4-formylphenyl)thiophene-2-yl),(CH₂)₅NHSO₂(5-pyrid-2-ylthiophene-2-yl),CH₂CH₂NHSO₂(5-(2-(morpholinoCH₂)phenyl)thiophene-2-yl),CH₂CH₂NHSO₂(5-(4-(morpholinoCH₂)phenyl)thiophene-2-yl),CH₂CH₂NHSO₂(5-(3-(piperidinylCH₂)phenyl)thiophene-2-yl),CH₂CH₂NHSO₂-(5-(2-formylphenyl)thiophene-2-yl), CH₂CH₂NHSO₂(5-(3-(Et₂NCH₂) phenyl) thiophene-2-yl, CH₂CH₂NHSO₂ (5-(3-(Cyclohexyl(Me)NCH₂)phenyl)thiophene-2-yl), CH₂CH₂NHSO₂(5-(3-(pyrrolidinylCH₂)phenyl)thiophene-2-yl),CH₂CH₂NHSO₂(5-(3-cyanophenyl)thiophene-2-yl),CH₂CH₂NHSO₂(5-(3-(azetidinylCH₂)phenyl)thiophene-2-yl),CONHCH₂CH₂NHSO₂(5-(3-(N-ethyl-N-methylaminomethyl)phenyl)thiophene-2-yl),CH₂CH₂NHSO₂(5-(3-(bis(2-methoxyethyl)aminomethyl)phenyl)thiophene-2-yl),CH₂CH₂NHSO₂(5-(3-cyanophenyl)thiophene-2-yl), CH₂CH₂NHSO₂(5-(3-(thiomorpholinoCH₂)phenyl)thiophene-2-yl),CH₂CH₂NHSQ₂(5-(3-(4-ketopiperidinylCH₂)phenyl)thiophene-2-yl), Ms-L-Ser(Bn), and CH₂CH₂NHSO₂Ph.
 4. The compound of claim 2 wherein W, R₂ and Rare selected in accordance with the following table: W R₂ R  1 Ms-D- BnPh Ser(Bn)  2 PhSO₂-L- Bn Ph Pro  3 Cbz-L-Leu Bn Ph  4 Ms-D- Ser(Bn) Bn

 5 Ms-D- Ser(Bn) Bn

 6 PhSO₂-L- Bn Ph Phe  7 Ms-D- Ser(Bn) Bn

 8 Ms-D- Ser(Bn) Bn

 9 Ms-D- Ser(Bn) Bn

10 Ms-D- Ser(Bn) CH₂OMe

11 Ms-D- Ser(Bn) Bn

12 Ms-D- CH₂OMe Ph Ser(Bn) 13 Ms-D- Ser(Bn) Bn

14 Ms-D- Ser(Bn) Bn

15 Ms-D- Ser(Bn) Bn

16 Ms-D- Ser(Bn) Bn

17 Ms-D- Ser(Bn) Bn

18 Ms-D- Ser(Bn) Bn


5. The compound of claim 3 wherein W and R are selected in accordancewith the following table: W R  1 Ms-D-SerCH₂CH₂NHSO₂-(3-(2-NH₂-thiazol-4-yl)Ph) (Bn)  2 Ms-D-SerCH₂CH₂NHSO₂-(5-(3-formylphenyl)thiophene-2-yl) (Bn)  3 Ms-D-SerCH₂CH₂NHSO₂(5-(3-(Me₂NCH₂)phenyl)thiophene- (Bn) 2-yl)  4 Ms-D-SerCH₂CH₂NHSO₂(5-(3-(morpholinoCH₂)phenyl) (Bn) thiophene-2-yl)  5 Ms-D-SerCH₂CH₂NHSO₂(4-(MorpholinoCH₂)Ph) (Bn)  6 Ms-D-SerCH₂CH₂NHSO₂(5-(3-(N-Me-piperazinyl- (Bn) CH₂)phenyl)thiophene-2-yl)  7Ms-D-Ser CH₂CH₂NHSO₂(5-(3-(HOCH₂)phenyl)thiophene-2-yl) (Bn)  8 Ms-D-SerCH₂CH₂SO₂NHPh (Bn)  9 Ms-D-Ser CH₂CH₂SO₂NH(4 -CF₃Ph) (Bn) 10 Ms-D-SerCH₂CH₂N(Me)SO₂-(5-(3-formylphenyl)thiophene- (Bn) 2-yl) 11 Ms-D-ThrCH₂CH₂NHSO₂(5-pyrid-2-ylthiophene-2-yl) (Bn) 12 Ms-(D, L)-CH₂CH₂NHSO₂(5-pyrid-2-ylthiophene-2-yl) Phenylgly 13 Ms-(D, L)-CH₂CH₂NHSO₂Ph Phenylgly 14 Ms-D-Thr CH₂CH₂NHSO₂(5-(3- (Bn)(morpholinoCH₂)phenyl)thiophene-2-yl) 15 Ms-D-PheCH₂CH₂NHSO₂(5-pyrid-2-ylthiophene-2-yl) 16 Ms-D-SerCH₂CH₂NHSO₂(5-(3-Fluorophenyl)thiophene-2-yl) (Bn) 17 Ms-D-SerCH₂CH₂NHSO₂-(5-(3-Nitrophenyl)thiophene-2-yl) (Bn) 18 Ms-D-SerCH₂CH₂NHSO₂-(5-(3-Methylphenyl)thiophene-2-yl) (Bn) 19 Ms-D-SerCH₂CH₂NHSO₂(5-(3-(AcNH)phenyl)thiophene-2-yl) (Bn) 20 Ms-D-SerCH₂CH₂NHSO₂(5-(3-(CH₃CO)phenyl)thiophene-2-yl) (Bn) 21 Ms-D-SerCH₂CH₂NHSO₂-(5-(4-Methoxyphenyl)thiophene- (Bn) 2-yl) 22 Cbz-Leu-CH₂CH₂NHSO₂(5-pyrid-2-ylthiophene-2-yl) Leu 23 Ms-D-SerCH₂CH₂NHSO₂-(5-(4-formylphenyl)thiophene- (Bn) 2-yl) 24 Ms-D-SerCH₂CH₂NHSO₂(4-(2- (Bn) (MeOCH₂)PyrrolidinylCH₂)Ph) 25 Ms-D-SerCH₂CH₂NHSO₂(5-(2- (Bn) (morpholinoCH₂)phenyl)thiophene-2-yl) 26 Ms-D-SerCH₂CH₂NHSO₂(5-(4- (Bn) (morpholinoCH₂)phenyl)thiophene-2-yl) 27 Ms-D-SerCH₂CH₂NHSO₂(5-(3- (Bn) (piperidinylCH₂)phenyl)thiophene-2-yl) 28Ms-D-Ser CH₂CH₂NHSO₂-(5-(2-formylphenyl)thiophene- (Bn) 2-yl) 29Ms-D-Ser CH₂CH₂NHSO₂(5-(3- (Bn) (Et₂NCH₂)phenyl)thiophene-2-yl) 30Ms-D-Ser CH₂CH₂NHSO₂(5-(3- (Bn) (Cyclohexyl(Me)NCH₂)phenyl)thiophene-2-yl) 31 Ms-D-Ser CH₂CH₂NHSO₂(5-(3- (Bn)(pyrrolidinylCH₂)phenyl)thiophene-2-yl) 32 Ms-D-SerCH₂CH₂NHSO₂(5-(3-cyanophenyl)thiophene-2- (Bn) yl) 33 Ms-D-SerCH₂CH₂NHSO₂(5-(3- (Bn) (azetidinylCH₂)phenyl)thiophene-2-yl) 34 Ms-D-SerCONHCH₂CH₂NHSO₂(5-(3-(N-ethyl-N- (Bn)methylaminomethyl)phenyl)thiophene-2-yl) 35 Ms-D-SerCH₂CH₂NHSO₂(5-(3-(bis(2- (Bn)methoxyethyl)aminomethyl)phenyl)thiophene-2-yl) 36 Ms-D-SerCH₂CH₂NHSO₂(5-(3-cyanophenyl)thiophene-2-yl) (Bn) 37 Ms-D-SerCH₂CH₂NHSO₂(5-(3- (Bn) (thiomorpholinoCH₂)phenyl)thiophene-2-yl) 38Ms-D-Ser CH₂CH₂NHSO₂(5-(3-(4- (Bn)ketopiperidinylCH₂)phenyl)thiophene-2-yl) 39 Ms-L-Ser CH₂CH₂NHSO₂Ph(Bn).